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Introduction
The Widal test is an advanced way to check for antibodies that your
body makes against the salmonella bacteria that causes typhoid
fever. It looks for O and H antibodies in a patient’s sample blood
(serum).
Keep reading this blog to understand the meaning of the Widal test,
its principle, procedure, and test result interpretation.
On the other hand, the bacteria S. Paratyphi has the following two
antigens:
1. S. Paratyphi A
2. S. Paratyphi B
This agglutination test detects the antibodies your body has made in
response to a particular bacteria or antigen. If you have typhoid
fever, your sera (blood) will possess antibodies that will react and
agglutinate salmonella antigens in an agglutination test.
The following step of this test measures the titre of the positive
antigen:
1. Patient’s serum
2. Pipette (lab tool)
3. Serum
4. S. Antigen ( O, H, AH, BH)
5. Slide
6. Mix Stick
7. Stopwatch
For this test, you will use a slide with 6 reaction circles, marked as O,
H, AH, BH, PC and NC. To begin with,
1. Put one drop of the patient’s serum in four reaction circles, i.e.,
O, H, AH, BH.
2. Add one drop of positive control in the PC circle and one in the
NC circle.
3. Next, add one drop of O antigen in the O circle, P antigen in the
P circle, AH antigen in the AH circle, and BH antigen in the BH
circle, respectively.
4. Add any antigen, i.e., O, H, AH, BH in both PC and NC.
5. Next, mix the serum and antigen in each circle properly so that
the mixture doesn’t go out of the circle and touch the slide.
6. Also, one mixture should not mix with another, as it can
influence the test results.
7. Finally, rotate the slide in a slow circular motion to ensure a
proper mixture of serum and reagent.
Once everything is done, you can see the results. If the test is
positive, the test will be similar to the PC (+ve control circle), and if
it’s negative, it will be similar to NC (-ve control circle). In other
words, if there is any agglutination, the test results will be positive
and vice versa.
Now, if the test results are positive, the next stage will involve a
quantitative test. To confirm the diagnosis of typhoid fever, we will
take the reagent of the antigen that is positive. For example, if O was
positive in the qualitative test, we will take the O reagent in the
quantitative test.
1. Use a different slide with 8 circles: four for O antigen and four
for H.
2. Now, if O was positive in the last test, put 5 ul patient’s serum in
the 1st O circle, 10 ul in the 2nd, 20 ul in the 3rd, and 40 ul in the
4th circle horizontally.
3. In the same way, put one drop of the specific reagent in all four
circles.
4. To report, mark the values from the right side. Mark 1:40 on the
fourth circle, 1: 80 on the 3rd circle, 1:160 on the 2nd, and 1:320
on the 1st circle.
After mixing the serum and reagent properly and rotating the slide,
wait for the results to show. The result will be positive if it shows
positive in more than 100 in the O circle and 200 in the case of H.
1. Patient’s serum
2. O, H, AH and BH Antigens
3. Normal Saline
4. Pipette
5. Test Tube Rack
6. Test Tubes
7. Water Bath
Before beginning this test, it is essential to note that the tube method
is a dilution technique that has to be done correctly. Let’s understand
:
this test in a simpler language.
1. First, take nine tubes and arrange them in the rack. In the case
of O, mark the tubes in numbers from 1 to 9.
2. Add and mix 0.1 ml normal saline and 0.9 ml serum in the first
test tube. On the other hand, add 0.5 normal salines to each
remaining tube.
3. Next, take 0.5 ml from the 1st tube and add in the second. This
will result in 0.5 ml remaining solution in test tube 1 and 1 ml in
test tube 2nd.
4. Repeat this process i.e., take 0.5 ml from the last tube and add it
to the next tube to make it 1 ml. With the 8th tube, take 0.5 ml
and keep that in another separate tube.
5. Mix all the tubes properly. This will give us primary serial dilution
of all the tubes from 1st to 8th as 1:10, 1:20, 1:40, 1:80, 1:160,
1:320, 1:640, and 1:1280, respectively.
6. Take a new (9th) test tube and add positive control.
7. Next, add 0.5 ml of respective antigen (O, H, AH, BH) in all eight
tubes. This will make the final volume of each tube 1 ml.
8. After adding a reagent to all the tubes, we will have the final
serial dilution of all the tubes from 1st to 8th as 1:20, 1:40, 1:80,
1:160, 1:320, 1:640, 1:1280, 1:2580.
9. Now, mix well, cover and incubate the tubes at 37° C overnight
(18 to 24 hours).
If you have typhoid fever and there is some agglutination, you will
see that the 9th tube (positive control) will look similar to one of the
eight other tubes. If there is no enteric fever, there will be no change
in the normal range of the eight tubes and widal test.
If the tube that showed agglutination has a titre of more than 1:100 in
:
case of O and 1: 200 in H, it will be widal test positive (active
infection). Other than this, rest titers are considered the normal
range of a widal test.
1. The results of the Widal test can be falsely positive in the case
of past vaccination or S. Typhi infection.
2. The Widal test is time-consuming; until a diagnosis is made, it
becomes too late to start the treatment.
3. A widal test can not distinguish between a patient’s past
infection, current infection, or a S. Typhi vaccination.
4. The test results can be falsely positive in typhus, acute
falciparum malaria, chronic liver disease, rheumatoid arthritis,
nephrotic syndrome and myelomatosis.
5. Because so many factors can influence the test results, it is
better to not just depend on this test for typhoid diagnosis.
Conclusion
If you have typhoid fever or are experiencing its symptoms, you can
book a Widal test today.
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